Mutational analysis of IDH1 codon 132 in glioblastomas and other common cancers

Missense somatic mutations in IDH1 gene affecting codon 132 have recently been reported in glioblastoma multiforme (GBM) and other gliomas. The recurrent nature of the IDH1 mutations in the same amino acid strongly suggests that the mutations may play important roles in the pathogenesis of glial tumors. The aim of this study was to see whether the IDH1 codon 132 mutations occur in other human cancers besides glial tumors. We also attempted to confirm the occurrence of the IDH1 mutations in GBM of Korean patients. We have analyzed 1,186 cancer tissues from various origins, including carcinomas from breast, colon, lung, stomach, esophagus, liver, prostate, urinary bladder, ovary, uterine cervix, skin and kidney, and malignant mesotheliomas, primary GBM, malignant meningiomas, multiple myelomas and acute leukemias by single‐strand conformation polymorphism analysis. We found four IDH1 codon 132 mutations in the GBM (4/25; 16.0%), two in the prostate carcinomas (2/75; 2.7%) and one in the B‐acute lymphoblastic leukemias (B‐ALL) (1/60; 1.7%), but none in other cancers. The IDH1 mutations consisted of five p.R132H and two p.R132C mutations. The data indicate that IDH1 codon 132 mutations occur not only in GBM, but also in prostate cancers and B‐ALL. This study suggests that despite the infrequent incidence of the IDH1 mutations in prostate cancers and B‐ALL, mutated IDH1 could be therapeutically targeted in these cancers and in glial tumors with the IDH1 mutations. © 2009 UICC     

Molecular Investigation of Isocitrate Dehydrogenase Gene (IDH) Mutations in Gliomas: First Report of IDH2 Mutations in Indian Patients

Recent genome wide sequencing has identified mutations in IDH1/IDH2 predominantly in grade II-III gliomasand secondary glioblastomas which are associated with favorable clinical outcome. These mutations have becomemolecular markers of significant diagnostic and prognostic relevance in the assessment of human gliomas. In thecurrent study we evaluated IDH1 (R132) and IDH2 (R172) in 32 gliomas of various grades and tumor subtypes.Sequencing analysis revealed R132H mutations in 18.7% tumors, while none of the cases showed IDH2 (R172)mutations. The frequency of IDH1 mutations was higher in females (21.4%) than males (11.1%), and it wassignificantly higher in younger patients. Histological analyses demonstrated presence of necrosis and microvascular proliferation in 69% and 75% respectively. Interestingly, IDH1 mutations were predominantly presentin non-necrotic tumors as well as in cases showing microvascular proliferation. Of the six IDH1 positive cases,three were glioblastomas (IV), and one each were anaplastic oligoastrocytoma (III), anaplastic oligodendrogliomaIII (n=1) and diffuse astrocytoma. In conclusion, IDH1 mutations are quite frequent in Indian glioma patientswhile IDH2 mutations are not observed. Since IDH mutations are associated with good prognosis, their use inroutine clinical practice will enable better risk stratification and management of glioma patients.     

Zonula Occludens-1, Occludin, and E-cadherin Protein Expression in Biliary Tract Cancers

The incidence of cholangiocarcinomas originating from intrahepatic and extrahepatic bile ducts, as well as of gallbladder carcinoma is increasing worldwide. The malignant transformation of biliary epithelia involves profound alterations of proteins in the intercellular junctions, among others zonula occludens-1 (ZO-1), occludin, and E-cadherin. Each plays important role in the maintenance of epithelial cell polarity and regulation of cell growth and differentiation. Our aim was to investigate ZO-1, occludin, and E-cadherin immunohistochemical reactions in tissue microarray blocks containing 57 normal and 62 neoplastic biliary tract samples. We demonstrated that the tight junction components ZO-1, occludin, and E-cadherin are downregulated in carcinomas arising from various compartments of the biliary tract (normal intrahepatic and extrahepatic bile ducts, gallbladder) as compared with their normal sites of origin. These results were confirmed by discriminant analysis yielding clear separation of the three normal sample groups from carcinomas in the corresponding locations.     

Multigene mutational profiling of cholangiocarcinomas identifies actionable molecular subgroups

One-hundred-fifty-three biliary cancers, including 70 intrahepatic cholangiocarcinomas (ICC), 57 extrahepatic cholangiocarcinomas (ECC) and 26 gallbladder carcinomas (GBC) were assessed for mutations in 56 genes using multigene next-generation sequencing. Expression of EGFR and mTOR pathway genes was investigated by immunohistochemistry. At least one mutated gene was observed in 118/153 (77%) cancers. The genes most frequently involved were KRAS (28%), TP53 (18%), ARID1A (12%), IDH1/2 (9%), PBRM1 (9%), BAP1 (7%), and PIK3CA (7%). IDH1/2 (p=0.0005) and BAP1 (p=0.0097) mutations were characteristic of ICC, while KRAS (p=0.0019) and TP53 (p=0.0019) were more frequent in ECC and GBC. Multivariate analysis identified tumour stage and TP53 mutations as independent predictors of survival. Alterations in chromatin remodeling genes (ARID1A, BAP1, PBRM1, SMARCB1) were seen in 31% of cases. Potentially actionable mutations were seen in 104/153 (68%) cancers: i) KRAS/NRAS/BRAF mutations were found in 34% of cancers; ii) mTOR pathway activation was documented by immunohistochemistry in 51% of cases and by mutations in mTOR pathway genes in 19% of cancers; iii) TGF-ß/Smad signaling was altered in 10.5% cancers; iv) mutations in tyrosine kinase receptors were found in 9% cases. Our study identified molecular subgroups of cholangiocarcinomas that can be explored for specific drug targeting in clinical trials.     

Alterations of p16 and prognosis in biliary tract cancers from a population-based study in China.

PURPOSE: Biliary tract cancer is an uncommon malignancy with a poor survival rate. We evaluated p16 gene alteration as a prognostic marker for this disease. EXPERIMENTAL DESIGN: We studied p16 gene alterations by sequencing, methylation, and loss of heterozygosity of chromosome 9p in 118 biliary tract carcinomas, including 68 gallbladder cancers, 33 extrahepatic bile duct cancers, and 17 ampullary cancers. Survival was evaluated in 57 patients with gallbladder carcinomas, 27 with bile duct carcinomas, and 16 with ampullary carcinomas with and without somatic p16 alterations detected by two different methods. RESULTS: p16 gene alterations including silent mutations were present in 61.8% gallbladder cancers, 54.5% bile duct cancers, and 70.6% ampullary cancers. p16 gene nonsilent mutations, p16 methylation, and loss of chromosome 9p21-22 that targets p14, p15, and p16 genes were present in 13 of 53 (24.5%), 8 of 54 (14.8%), and 32 of 44 (72.7%) gallbladder tumors; 5 of 25 (20.0%), 5 of 31 (16.1%), and 12 of 21 (57.1%) bile duct tumors; and 3 of 13 (23.1%), 6 of 15 (40.0%), and 8 of 16 (50.0%) ampullary tumors, respectively. The mean survival of patients with gallbladder cancers without p16 alterations was 21.5 +/- 14.8 months compared with 12.1 +/- 11.4 months for patients with p16 alterations (P = 0.02). CONCLUSIONS: Alteration of p16 gene alone or in combination with alterations of other tumor suppressor genes on chromosome 9p is a prognostic indicator in gallbladder carcinoma, with more favorable survival rates associated with carcinomas lacking p16 gene alterations.     

血液肿瘤患者IDH1/IDH2基因突变发生率及其血液病分布特点研究

目的研究血液肿瘤患者IDH1/IDH2基因突变发生率和其血液病的分布特点。方法选取300例不同病情的血液患者骨髓的样品,其中245例是血液肿瘤患者,55例是非血液肿瘤患者,对比检查IDH2和IDH1基因是否在血液肿瘤患者中有突变行为。结果实验共发现了7个错义的突变基因,其中2个为IDH2基因突变,4个为IDH1基因突变;还有2个IDH1同义突变。4个IDH1的突变类型是:199M、R132C、R131D及R132H,前3个突变患者都出现在3例急性髓系的白血病患者中,最后1例出现在急性淋巴细胞的白血病患者中。2个IDH2的错义突变类型是:P167R和R172K,这2例也都出现在急性髓系的白血病患者中。在发生IDH1错义基因突变的4例患者中,其体内白细胞的计数明显升高,且患者年龄均大于60岁。在发生IDH2基因突变的2例患者中,其体内白细胞的计数明显降低,且患者的年龄均小于60岁。结论 96例的急性白血病患者的检测中,共发现了9例突变患者,其IDH的突变率是9.3%,且7例发生于急性髓系的白血病患者中,说明急性髓系的白血病的发病和IDH的突变有紧密的联系。     

Mutant IDH1 is required for IDH1 mutated tumor cell growth

Frequent somatic hotspot mutations in isocitrate dehydrogenase 1 (IDH1) have been identified in gliomas, acute myeloid leukemias, chondrosarcomas, and other cancers, providing a likely avenue for targeted cancer therapy. However, whether mutant IDH1 protein is required for maintaining IDH1 mutated tumor cell growth remains unknown. Here, using a genetically engineered inducible system, we report that selective suppression of endogenous mutant IDH1 expression in HT1080, a fibrosarcoma cell line with a native IDH1R132C heterozygous mutation, significantly inhibits cell proliferation and decreases clonogenic potential. Our findings offer insights into changes that may contribute to the inhibition of cell proliferation and offer a strong preclinical rationale for utilizing mutant IDH1 as a valid therapeutic target.     

Matching genomic molecular aberrations with molecular targeted agents: Are biliary tract cancers an ideal playground?

Biliary tract cancers (BTCs) are a heterogeneous group of tumours with geographical discrepancies in terms of incidence and risk factors. However, a convergent genomic and epigenetic mutational landscape emerges from the genome-wide screens of BTCs in South East Asia, Latin America and in the Western World. Specificities are observed for some alterations and anatomical subtypes: frequent fibroblast growth factor receptor 2 (FGFR2) and isocitrate dehydrogenase 1/2 (IDH1/2) alterations are specific to intrahepatic cholangiocarcinomas (ICCs), whereas frequent ERBB2 oncogene alterations are specific to extrahepatic cholangiocarcinomas (ECCs) and gallbladder carcinomas (GBCs). Until now, the outcome of patients with BTCs treated by molecular targeted agents (MTAs) alone or in combination with conventional chemotherapy in non-biology driven trials remains poor and does not exceed the outcome of patients treated with chemotherapy alone. Encouraging reports of biology-driven therapeutic approaches should accelerate the clinical development of MTAs in BTCs. Additionally, frequent epigenetic aberrations such as IDH1/2 mutations and switch/sucrose non-fermenting (SWI/SNF) complex dysfunctions suggest that epidrugs must also be considered. In this review, we expose the rationale and feasibility to biologically drive the treatment of BTC patients.     

Mutation Analysis of IDH1/2 Genes in Unselected De novo Acute Myeloid Leukaemia Patients in India - Identification of A Novel IDH2 Mutation

IDH1/2 mutations which result in alternation in DNA methylation pattern are one of the most commonmethylation associated mutations in Acute myeloid leukaemia. IDH1/2 mutations frequently associated withhigher platelet level, normal cytogentics and NPM1 mutations. Here we analyzed IDH1/2 mutations in 200 newlydiagnosed unselected Indian adult AML patients and investigated their correlation with clinical, cytogeneticparameters along with cooperating NPM1 mutation. We detected 5.5% and 4% mutations in IDH1/2 genes,respectively. Except IDH2 c.515_516GG>AA mutation, all the other identified mutations were reported mutations.Similar to reported c.515G>A mutation, the novel c.515_516GG>AA mutation replaces 172nd arginine to lysinein the active site of the enzyme. Even though there was a preponderance of IDH1/2 mutations in NK-AML,cytogenetically abnormal patients also harboured IDH1/2 mutations. IDH1 mutations showed significant higherplatelet count and NPM1 mutations. IDH2 mutated patients displayed infrequent NPM1 mutations and lowerWBC count. All the NPM1 mutations in the IDH1/2 mutated cases showed type A mutation. The present datasuggest that IDH1/2 mutations are associated with normal cytogenetics and type A NPM1 mutations in adultIndian AML patients.     

Association of The IDH1 C.395G>A (R132H) Mutation with Histological Type in Malay Brain Tumors

Background: Brain tumors, constituting one of the most deadly forms of cancer worldwide, result from the accumulation of multiple genetic and epigenetic alterations in genes and signaling pathways. Isocitrate dehydrogenase enzyme isoform 1 (IDH1) mutations are frequently identified in primary brain tumors and acute myeloid leukemia. Studies on IDH1 gene mutations have been extensively performed in various populations worldwide but not in Malaysia. This work was conducted to study the prevalence of IDH1 c.395G>A (R132H) hotspot mutations in a group of Malaysian patients with brain tumors in order to gain local data for the IDH1 mutation profile in our population. Methods: Mutation analysis of c.395G>A (R132H) of IDH1 was performed in 40 brain tumor specimens by the polymerase chain reaction-restriction fragment length polymorphism method (PCR-RFLP) and then verified by direct sequencing. Associations between the IDH1 c.395G>A (R132H) mutation and clinicopathologic characteristics were also analyzed. Results: The IDH1 c.395G>A (R132H) mutation was detected in 14/40 patients (35%). A significant association was found with histological tumor types, but not with age, gender and race. Conclusions: IDH1 is frequently mutated and associated with histological subtypes in Malay brain tumors.     

Beta-catenin mutations in biliary tract cancers: a population-based study in China

beta-Catenin is an ubiquitously expressed cytoplasmic protein that has a crucial role in both cadherin-mediated cell-cell adhesion and as a downstream signaling molecule in the wingless/Wnt pathway. Activating mutations in exon 3 of the beta-catenin gene, at the phosphorylation sites for ubiquitination and degradation of beta-catenin, are present in a variety of cancers. Because alterations of the adenomatous polyposis coli (APC) gene are present in biliary tract cancers and the APC protein modulates levels of beta-catenin, we evaluated the role of beta-catenin in biliary tract cancer by sequencing the third exon of the beta-catenin gene among 107 biliary tract cancers and 7 gallbladder adenomas from a population-based study in CHINA: Point mutations of serine or threonine phosphorylation sites in exon 3 of beta-catenin were present in 8 of 107 (7.5%) biliary tract cancers and 4 of 7 (57.1%) gallbladder adenomas. Mutations of beta-catenin were more frequent in ampullary and gallbladder carcinomas than in bile duct carcinomas (P = 0.04) and in papillary adenocarcinomas than other histological types of carcinomas (P = 0.02). These results suggest that the molecular pathways of biliary tract neoplasms vary by anatomical subsite and histological subtype.     

皮肤黑色素瘤中IDH1基因突变分析

目的:应用癌症基因组图谱(TCGA)数据库分析皮肤黑色素瘤中IDH1基因突变情况.方法:从癌症基因组图谱(TCGA)数据库收集皮肤黑色素瘤数据集,统计患者临床信息、应用cBioPortal网站对数据集进行突变、离散基因、基因共表达、生存期等分析.结果:287个病例中,IDH1基因突变15例,其中,R132C突变10例,R132L突变2例,P33S/E361K突变各1例,X174缺失1例.结论:IDH1基因突变在皮肤黑色素瘤中有一定发生率,占比5.2%,且以R132突变为主,此位点有可能成为靶向药物作用的靶点.     

Trends in the incidence of primary liver and biliary tract cancers in England and Wales 1971-2001

In the last two decades, mortality from primary liver cancer has increased in the UK. We aimed to determine whether the incidence trends for these cancers were similar and in particular if the increasing occurrence of cholangiocarcinoma has continued. We calculated directly age-standardised incidence rates (using the European standard population) by subsite and histological type for all cancers of the liver, gallbladder and biliary tract in England and Wales from 1971 to 2001, using cancer registry data. The incidence of cancers of the liver, gallbladder and biliary tract increased, with the greatest rise, around 12-fold, in intrahepatic bile duct cancers. The rate of liver cell cancer increased by around 45% in males, but by <10% in females. There were marked reductions in the incidence of gallbladder and extrahepatic bile duct cancer. Cholangiocarcinoma increased around 16-fold and became the most common type of primary liver cancer in females, while hepatocellular carcinoma remained the commonest type in males. The age-specific incidence rates showed that intrahepatic bile duct cancer continued to increase throughout the 1990s in those aged 75 and over, while liver cell cancer decreased in the older age groups. In conclusion, there were increases in the incidence of primary liver cancer, which have been particularly dramatic for intrahepatic bile duct cancer, over the last three decades of the 20th century in England and Wales. There has been a halving in the incidence of gallbladder cancer and a reduction of a third in extrahepatic bile duct cancer.     

Radiotherapy and razoxane in advanced bile duct carcinomas

OBJECTIVES: Little is known about the radiation sensitivity of bile duct carcinomas. The current study was undertaken to prospectively assess the objective response rates in bile duct carcinomas treated with radiotherapy and razoxane. MATERIALS AND METHODS: Twenty-three patients with advanced cancer of the biliary tree were irradiated together with the radiosensitizer razoxane at a dose of 125 mg twice daily by mouth. There were 16 females and 7 males, median age 68 years. They received a total tumor dose of 48 Gy (range 1.7-60) at the ICRU point with single fractions of 1.7 to 2 Gy. RESULTS: Among the 23 patients, 14 had measurable disease. Objective tumor responses were seen in 4/4 gallbladder carcinomas (1 CR, 3 PR), 4/5 extrahepatic cholangiocarcinomas (2 CR 2 PR), and 1/5 hepatobiliary cancers (1 PR), leading to an overall response rate of 64%. The tumors remained locally controlled in 12 out of 16 assessable patients (75%). On an intention-to-treat basis, all patients with different biliary cancer without distant metastases had a median-survival time of 10 months (range 1 to 48) from the start of the radiotherapy; the 1-year survival was 43%. No patient survived beyond 4 years. Tolerance to the treatment was fairly good. Nausea and vomiting of grade 1 and 2 (WHO) was noted in 61%, and reversible leukopenia of grade 3 and 4 in 9% of the cases. The rate of inherent complications was high. CONCLUSION: Combined radiotherapy and razoxane led to local response rates which are superior to data from the literature when radiotherapy alone is used. Obstacles to the treatment were complications of the disease and frequent metastasis.     

Prognostic value of IDH1 mutations identified with PCR-RFLP assay in acute myeloid leukemia patients

BackgroundSomatic mutations in isocitrate dehydrogenase 1 (IDH1) gene occur frequently in primary brain tumors. Recently theses mutations were demonstrated in acute myeloid leukemia (AML). So far, assessment of these mutations relied on the DNA sequencing technique.Aim of the workThe aim of this study was to detect somatic mutations in IDH1 gene using mismatched primers suitable for endonuclease based detection, without the need for DNA sequencing, and to estimate its prognostic value, on patients with de novo AML.MethodsResidual DNA extracted from pretreatment bone marrow (BM) samples of 100 patients with de novo AML was used. The polymerase chain reaction-restriction fragment length polymorphism method (PCR-RFLP) was adapted to IDH1gene, codon 132 mutations screening.ResultsThe frequency of IDH1 mutations was 13%. In the non-acute promyelocytic leukemia group (non-APL), IDH1 mutations were significantly associated with FLT3-ITD negative patients (p = 0.03). Patients with IDH1 mutations did not achieve complete remission (CR). There was a trend for shorter overall survival (OS) in patients with IDH1 mutation compared to those with wild type (p = 0.08).ConclusionIDH1 mutations are recurring genetic alterations in AML and they may have unfavorable impact on clinical outcome in adult AML. The PCR-RFLP method allows for a fast, inexpensive, and sensitive method for the detection of IDH1 mutations in AML.     

Analysis of IDH1 and IDH2 mutations in Japanese glioma patients

A recent study reported on mutations in the active site of the isocitrate dehydrogenase 1 ( IDH1 ) gene in several types of gliomas. All mutations detected resulted in an amino acid exchange at position 132. We analyzed the genomic region spanning wild-type R132 of IDH1 by direct sequencing in 125 glial tumors. A total of 39 IDH1 mutations were observed. Mutations of the IDH2 gene, homologous to IDH1 , were often detected in gliomas without IDH1 mutations. In the present study, R172 mutation of the IDH2 gene was detected in one anaplastic astrocytoma. IDH1 or IDH2 mutations were frequently in oligodendrogliomas (67%), anaplastic astrocytomas (62%), anaplastic oligoastrocytomas (75%), anaplastic oligodendrogliomas (50%), secondary glioblastomas (67%), gangliogliomas (38%), and anaplastic gangliogliomas (60%). Primary glioblastomas were characterized by a low frequency of mutations (5%) at amino acid position 132 of IDH1 . Mutations of the IDH1 or IDH2 genes were significantly associated with improved outcome in patients with anaplastic astrocytomas. Our data suggest that IDH1 or IDH2 mutation plays a role in early tumor progression of several types of glioma and might arise from a common glial precursor. The infrequency of IDH1 mutation in primary glioblastomas revealed that these subtypes are genetically distinct entities from other glial tumors. ( Cancer Sci  2009; 100: 1996–1998)     

Molecular signatures classify astrocytic gliomas by IDH1 mutation status

To identify novel glioma‐associated pathomechanisms and molecular markers, we performed an array‐based comparative genomic hybridization analysis of 131 diffuse astrocytic gliomas, including 87 primary glioblastomas (pGBIV), 13 secondary glioblastomas (sGBIV), 19 anaplastic astrocytomas (AAIII) and 12 diffuse astrocytomas (AII). All tumors were additionally screened for IDH1 and IDH2 mutations. Expression profiling was performed for 74 tumors (42 pGBIV, 11 sGBIV, 13 AAIII, 8 AII). Unsupervised and supervised bioinformatic analyses revealed distinct genomic and expression profiles separating pGBIV from the other entities. Classifier expression signatures were strongly associated with the IDH1 gene mutation status. Within pGBIV, the rare subtype of IDH1 mutant tumors shared expression profiles with IDH1 mutant sGBIV and was associated with longer overall survival compared with IDH1 wild‐type tumors. In patients with IDH1 wild‐type pGBIV, PDGFRA gain or amplification as well as 19q gain were associated with patient outcome. Array‐CGH analysis additionally revealed homozygous deletions of the FGFR2 gene at 10q26.13 in 2 pGBIV, with reduced FGFR2 mRNA levels being frequent in pGBIV and linked to poor outcome. In conclusion, we report that diffuse astrocytic gliomas can be separated into 2 major molecular groups with distinct genomic and mRNA profiles as well as IDH1 gene mutation status. In addition, our results suggest FGFR2 as a novel glioma‐associated candidate tumor suppressor gene on the long arm of chromosome 10.     

Frequent expression of the novel cancer testis antigen MAGE-C2/CT-10 in hepatocellular carcinoma

Cancer testis (CT) antigens are attractive targets for immunotherapy in cancer patients. Immunohistochemistry was used to study the expression of the CT antigens MAGE-C2/CT-10, MAGE-C1/CT-7, GAGE, MAGE-A4 and NY-ESO-1 in 146 hepatocellular carcinomas, 13 intrahepatic cholangiocarcinomas, 37 extrahepatic cholangiocarcinomas and 32 gallbladder carcinomas. Immunopositivity was correlated with clinicopathological parameters, MHC Class 1 expression, intratumoral CD4+, CD8+ and FOXP3+ T cells and CD163+ antigen-presenting cells. Of the 146 hepatocellular carcinomas, 34% were positive for MAGE-C2/CT-10, 12% for MAGE-C1/CT-7, 11% for GAGE and 2% for NY-ESO-1, respectively. MHC Class 1 coexpression was identified in almost all CT antigen-positive tumors. The number of intratumoral FOXP3+ regulatory T cells was increased in CT antigen-positive hepatocellular carcinomas (p<0.004), suggesting inhibition of immune response in such tumors. Furthermore, MAGE-C1/CT-7 and GAGE positivity was correlated with reduced overall survival in patients with hepatocellular carcinoma (p=0.03 and 0.01, respectively). Four (13%) gallbladder carcinomas stained positive for MAGE-C2/CT-10, of which 1 tumor (3%) was also positive for NY-ESO-1 and GAGE. CT antigens were not expressed in intra- and extrahepatic cholangiocarcinomas. Our results suggest that MAGE-C2/CT-10 may be a good candidate for peptide vaccination in patients with hepatocellular carcinoma.     

K-ras mutation, p53 overexpression, and microsatellite instability in biliary tract cancers: a population-based study in China.

PURPOSE: The genetic alterations in biliary tract cancer and clinicopathological associations have not been studied in large population-based studies. Experimental Design: We evaluated genetic alterations such as K-ras mutation, p53 overexpression, microsatellite instability (MSI), and alterations of the polyadenine tract present in the transforming growth factor beta receptor type II (TGFbetaRII) gene in 126 biliary tract cancers: 75 gallbladder cancers, 33 bile duct cancers, and 18 ampullary cancers. These genetic alterations were compared with patient demographics and clinicopathological characteristics of the tumors. RESULTS: Mutation of the K-ras gene was present in 18 of 126 (14.3%) biliary tract cancers. K-ras mutation was present in 11 of 18 (61.1%) ampullary cancers, 5 of 33 (15.2%) bile duct cancers, and 2 of 75 (2.7%) gallbladder cancers (P = 0.000001). The mean survival of patients who had bile duct carcinomas with K-ras mutation was 3.0 +/- 2.2 months compared with 15.5 +/- 12.5 months for those without mutation (P = 0.03) but was not different for other tumor sites. p53 overexpression was present in 34 of 123 (27.6%) cancers. MSI-high (allelic shifts in 40% or more loci or alteration of the TGFbetaRII gene) was present in 4 of 126 (3.2%) biliary tract cancers without hereditary nonpolyposis colorectal cancer. MSI-high was more common in mucinous adenocarcinomas (P = 0.006) and in patients with early age of onset of cancer (P = 0.04). CONCLUSIONS: The genetic alterations in biliary tract cancers are dependent on the tumor subsite, histology, and age of onset and are associated with prognosis.